Parameters of the reproductive tract, spermatogenesis, daily sperm production and major seminal plasma proteins of tropically adapted morada nova rams.

This study describes the reproductive parameters of Morada Nova rams, a breed of hair sheep from Brazil and with unique adaption to tropical environments. At 42 weeks of age, 15 rams were subjected to semen collection and, 1 week later, animals were slaughtered for collection of testes, epididymis and accessory sex glands. We conducted 2-D electrophoresis of seminal plasma proteins and major spots of stained gels were identified by LC-MS/MS. Total RNA was isolated from testis, epididymis and vesicular glands and subjected to qPCR. At slaughter, scrotal circumference and testicular weight were 27.5 ± 0.5 cm and 109.5 ± 6.0 g, respectively. Seminiferous tubule (ST) diameter was 188.3 ± 4.0 µm and each testis contained 1.9 ± 0.1 Sertoli cells (×10(9) ). Each Sertoli cell supported 0.1 ± 0.01 A spermatogonia, 3.0 ± 0.2 pachytene spermatocytes and 7.7 ± 0.5 round spermatids/tubule cross section. Daily sperm production reached 5.6 × 10(6)  cells/g of testis parenchyma. Testis size appeared as indicative of ST diameter and associated with epididymal measurements, as well as with the population of round spermatids and Sertoli cells/testis. Rams with heavier testes had greater daily sperm production and more Sertoli cells/testis. We detected 90.9 ± 9.6 spots per 2-D gel of seminal plasma. Major seminal proteins were identified as ram seminal vesicle proteins at 14 and 22 kDa, representing 16.2% and 12.8% of the total intensity of valid spots in the gels, respectively. Expression of both genes was greater in the vesicular glands as compared to testis and epididymis. Pixel intensity for those proteins in the 2-D gels was significantly correlated with seminal vesicle weight. This is the first description of the basic reproductive aspects of Morada Nova rams, including protein profiles of their seminal plasma. These findings will allow a better understanding of their reproductive physiology.

Topografia de ligação de proteínas do plasma seminal à membrana de espermatozoides bovinos epididimários e ejaculados / Binding patterns of seminal plasma plasma proteins on bovine epididymal and ejaculated sperm membrane

The present study was designed to investigate the topographical distribution of seminal plasma (SP) proteins on epididymal and ejaculated bovine sperm. Using immunocytochemistry and confocal microscopy the binding patterns of bovine SP proteins BSP-A3, albumin, transferrin, prostaglandin D-synthase (PGDS) and nucleobindin in ejaculated and cauda epididymal sperm from adult bulls were evaluated. Experiments were performed using sperm from 5 males. Data showed a positive signal, only detected for anti-PGDS, in the acrosomal cap of epididymal and ejaculated sperm. In ejaculated sperm, a very weak signal for nucleobindin 2 in the midpiece and equatorial regions was detected, using the anti-rat nucleobindin. BSP-A3 was detected on all sperm regions studied, with a more evidenced signal in acrosome and midpiece. However, no binding was detected for albumin or transferrin in neither epididymal nor ejaculated sperm. In conclusion, PGDS, BSP-A3 and nucleobindin interact directly with bovine sperm, with specific topographic distribution. These findings may add to the knowledge of how these proteins modulate sperm functions, thus providing fundamental support for studies designed to evaluate how they influence sperm functions.

Investigou-se a distribuição topográfica da ligação de proteínas seminais à membrana de espermatozoides bovinos epididimários e ejaculados. Utilizando imunocitoquímica e microscopia confocal, avaliaram-se a topografia de ligação das proteínas BSP-A3, albumina, transferrina, prostaglandina D sintetase (PGDS) e nucleobindina 2 (NUC2) à membrana espermática. Os experimentos foram realizados utilizando espermatozoides de cinco touros. Os resultados mostraram que, para espermatozoides epididimários, somente detectou-se a PGDS na crista do acrossomo. Nos espermatozoides ejaculados, a PGDS ligou-se de forma mais intensa à crista acrossômica, enquanto a NUC2 apresentou sinal bastante fraco na peça intermediária e região equatorial. A BSP-A3 ligou-se a todas as regiões estudadas, de forma mais intensa na peça intermediária e acrossomo. Nenhum sinal foi detectado para albumina ou transferrina, seja em espermatozoides epididimários ou ejaculados. Concluiu-se que PGDS, BSP-A3 e NUC2 interagem diretamente com espermatozoides bovinos, e mostrou distribuição topográfica específica. Estes achados permitem melhor compreensão sobre o papel desempenhado por essas proteínas na regulação da função espermática e da fertilidade.

Reproductive development of Santa Inês rams during the first year of life: body and testis growth, testosterone concentrations, sperm parameters, age at puberty and seminal plasma proteins.

We have investigated the reproductive development of the tropically adapted Santa Inês ram, the most common hair sheep in Brazil. From 8 to 48 weeks of age, 16 animals were evaluated for body and testis growth, semen parameters, testosterone concentrations and seminal plasma proteins, using two-dimensional SDS-PAGE. Animals were weaned at 30 days and kept in feedlots thereafter, receiving hay, concentrate (18% of crude protein) and mineral supplement. Body weight increased from 12.3 +/- 0.7 to 54.3 +/- 1.6 kg between 8 and 48 weeks (p < 0.05), but changes in thoracic perimeter and scrotal circumference were non-significant after 36 weeks (p > 0.05). The percentage of motile sperm increased slowly until 23 weeks and more rapidly after that age, but significant changes in progressive motility occurred after 25 weeks. Presence of abnormal sperm related inversely to age. Most significant changes in sperm concentration occurred between 38 and 44 weeks (0.38 +/- 0.05 to 1.14 +/- 0.24 x 10(9) cells/ml, p < 0.05) and testosterone reached its highest concentrations at 42 weeks, decreasing afterwards. Rams reached puberty at 28.2 +/- 0.8 weeks. The number of protein spots on seminal plasma gels was similar from 15 to 18 weeks (45 and 47 spots; p > 0.05), increased until 24 weeks (141 spots) and 28 weeks (170 spots; p < 0.05) and remained without significant (p > 0.05) changes from 28 to 48 weeks (186 +/- 10 spots). Furthermore, the intensity of selected spots on 2D maps increased (p < 0.05) between 15 and 28 weeks, which preceded or coincided with the main developmental changes in sperm motility and percentage of defective sperm in the ejaculates. These results will support future studies designed to characterize specific seminal plasma proteins whose expression relate to the development of testis, epididymis and accessory sex glands.